Cerebral small vessel disease predisposes to temporal lobe epilepsy in spontaneously hypertensive rats.

The link between cerebral small vessel disease (CSVD) and epilepsy has been poorly investigated. Some reports suggest that CSVD may predispose to temporal lobe epilepsy (TLE). Aim of this study was to evaluate whether spontaneously hypertensive rats (SHRs), an established model of systemic hypertension and CSVD, have a propensity to develop TLE more than generalized seizures. To this aim, amygdala kindling, as a model of TLE, and pentylenetetrazole (PTZ)-induced kindling, as a model of generalized seizures, have been used to ascertain whether SHRs are more prone to TLE as compared to Wistar Kyoto control rats. While young SHRs (without CSVD) do not differ from their age-matched controls in both models, old SHRs (with CSVD) develop stage 5 seizures in the amygdala kindling model (TLE) faster than age-matched control rats without CSVD. At odds, no differences between old SHRs and age-matched controls was observed in the development of PTZ kindling. Enalapril pre-treatment prevented the development of CSVD and normalized kindling development to control levels in SHRs. No difference was observed in the response to pharmacological treatment with carbamazepine or losartan. Overall, our study suggests that uncontrolled hypertension leading to CSVD might represent a risk factor for TLE. Further experimental studies are needed to unravel other risk factors that, along with CSVD, may predispose to TLE.

Anticonvulsive Activity in Audiogenic DBA/2 Mice of 1,4-Benzodiazepines and 1,5-Benzodiazepines with Different Activities at Cerebellar Granule Cell GABAA Receptors.

Herein, we tested in a model of generalized reflex epilepsy in mice different 1,4-benzodiazepines and 1,5-benzodiazepines with agonistic activity at the GABAA receptor population contributing to the peak component of the chloride current elicited by GABA in cerebellar granule cells (CGCs) in culture. The substances have all higher lipophilia than clobazam, an antiepileptic drug well known and used in human therapy. This ensures that they all can pass relatively easily the blood-brain barrier (BBB). The benzodiazepines were administered intraperitoneally (i.p.) and tested for their activity against sound-induced tonic and clonic seizures in a genetic model of experimental epilepsy, the DBA/2 mouse. Our data demonstrates an interesting inverse correlation between the ED50s and the efficacy (E %) of the drugs in increasing the peak chloride current elicited by GABA in cerebellar granule cells in culture. There is indication of the existence of a threshold of E % above which the increase of ED50 with increasing E % becomes linear. This is statistically significant for the clonic phase, whereas it is at the limit of significance for the tonic one. A possible interpretation of these results is that in this epilepsy model, projections from the cerebellum exert a convulsion prevention activity.

Unilateral and Bilateral Cortical Resection: Effects on Spike-Wave Discharges in a Genetic Absence Epilepsy Model.

RESEARCH QUESTION: Recent discoveries have challenged the traditional view that the thalamus is the primary source driving spike-and-wave discharges (SWDs). At odds, SWDs in genetic absence models have a cortical focal origin in the deep layers of the perioral region of the somatosensory cortex. The present study examines the effect of unilateral and bilateral surgical resection of the assumed focal cortical region on the occurrence of SWDs in anesthetized WAG/Rij rats, a well described and validated genetic absence model. METHODS: Male WAG/Rij rats were used: 9 in the resected and 6 in the control group. EEG recordings were made before and after craniectomy, after unilateral and after bilateral removal of the focal region. RESULTS: SWDs decreased after unilateral cortical resection, while SWDs were no longer noticed after bilateral resection. This was also the case when the resected areas were restricted to layers I-IV with layers V and VI intact. CONCLUSIONS: These results suggest that SWDs are completely abolished after bilateral removal of the focal region, most likely by interference with an intracortical columnar circuit. The evidence suggests that absence epilepsy is a network type of epilepsy since interference with only the local cortical network abolishes all seizures.

Statins and epilepsy: preclinical studies, clinical trials and statin-anticonvulsant drug interactions.

3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase inhibitors (statins) are potent cholesterol- lowering drugs which also possess beneficial antioxidant, antiinflammatory, immunomodulatory, and antiexcitotoxic effects. In addition, statins have proven neuroprotective effects in several neurological diseases: stroke, cerebral ischemia, Alzheimer's and Parkinson's disease, multiple sclerosis and traumatic brain injury. Relatively few studies have investigated the potential anti-seizure properties of statins in epilepsy and the possible underlying protective mechanisms that may be involved. This review summarizes the currently available data concerning statin effects in modulating seizure activity (sometimes adversely) and epileptogenesis in different experimental models as well as in clinical studies. Furthermore, we analyze the consequences of some of the more commonly reported statin-anticonvulsant drug interactions in the literature, discuss some of the adverse effects of statins encountered in clinical practice and comment on the potential future usefulness of statins in epilepsy therapy.

Skin rash during treatment with generic itraconazole.

Generic drugs have the same active substance, the same pharmaceutical form, the same therapeutic indications and a similar bioequivalence with the reference medicinal product (branded). Although a similar efficacy is postulated, some cases of clinical inefficacy during treatment with generic formulations have been reported. In this case, we describe a woman with onychomycosis that developed a skin rash during treatment with a generic formulation of itraconazole. Drug administration and its re-challenge confirmed the association between itraconazole and skin rash. Both Naranjo probability scale and World Health Organization causality assessment scale documented a probable association between generic-itraconazole and skin rash. The switch from generic formulation to brand one induced an improvement of symptoms. Since we are unable to evaluate the role of each excipient in the development of skin rash, we cannot rule out their involvement. However, more data are necessary to better define the similarities or differences between branded and generic formulations.

Hypericum perforatum: pharmacokinetic, mechanism of action, tolerability, and clinical drug-drug interactions.

Hypericum perforatum (HP) belongs to the Hypericaceae family and is one of the oldest used and most extensively investigated medicinal herbs. The medicinal form comprises the leaves and flowering tops of which the primary ingredients of interest are naphthodianthrones, xanthones, flavonoids, phloroglucinols (e.g. hyperforin), and hypericin. Although several constituents elicit pharmacological effects that are consistent with HP's antidepressant activity, no single mechanism of action underlying these effects has thus far been found. Various clinical trials have shown that HP has a comparable antidepressant efficacy as some currently used antidepressant drugs in the treatment of mild/moderate depression. Interestingly, low-hyperforin-content preparations are effective in the treatment of depression. Moreover, HP is also used to treat certain forms of anxiety. However, HP can induce various cytochrome P450s isozymes and/or P-glycoprotein, of which many drugs are substrates and which are the main origin of HP-drug interactions. Here, we analyse the existing evidence describing the clinical consequence of HP-drug interactions. Although some of the reported interactions are based on findings from in vitro studies, the clinical importance of which remain to be demonstrated, others are based on case reports where causality can, in some cases, be determined to reveal clinically significant interactions that suggest caution, consideration, and disclosure of potential interactions prior to informed use of HP.

Pharmacovigilance and drug safety in Calabria (Italy): 2012 adverse events analysis.

INTRODUCTION: Pharmacovigilance (PV) is designed to monitor drugs continuously after their commercialization, assessing and improving their safety profile. The main objective is to increase the spontaneous reporting of adverse drug reactions (ADRs), in order to have a wide variety of information. The Italian Drug Agency (Agenzia Italiana del Farmaco [AIFA]) is financing several projects to increase reporting. In Calabria, a PV information center has been created in 2010. MATERIALS AND METHODS: We obtained data using the database of the National Health Information System AIFA relatively to Italy and Calabria in the year 2012. Descriptive statistics were performed to analyze the ADRs. RESULTS: A total number of 461 ADRs have been reported in the year 2012 with an increase of 234% compared with 2011 (138 reports). Hospital doctors are the main source of this reporting (51.62%). Sorafenib (Nexavar(®)), the combination of amoxicillin/clavulanic acid and ketoprofen represent the drugs most frequently reported causing adverse reactions. Adverse events in female patients (61.83%) were more frequently reported, whereas the age groups "41-65" (39.07%) and "over 65" (27.9%) were the most affected. CONCLUSIONS: Calabria has had a positive increase in the number of ADRs reported, although it has not yet reached the gold standard set by World Health Organization (about 600 reports), the data have shown that PV culture is making inroads in this region and that PV projects stimulating and increasing PV knowledge are needed.

Limitations and obstacles of the spontaneous adverse drugs reactions reporting: Two "challenging" case reports.

INTRODUCTION: Nowadays, based on several epidemiological data, iatrogenic disease is an emerging public health problem, especially in industrialized countries. Adverse drugs reactions (ADRs) are extremely common and, therefore, clinically, socially, and economically worthy of attention. Spontaneous reporting system for suspected ADRs represents the cornerstone of the pharmacovigilance, because it allows rapid detection of potential alarm signals related to drugs use. However, spontaneous reporting system shows several limitations, which are mainly related to under-reporting. In this paper, we describe two particular case reports, which emphasize some reasons of under-reporting and other common criticisms of spontaneous reporting systems. MATERIALS AND METHODS: We performed a computer-aided search of Medline, PubMed, Embase, Cochrane library databases, national and international databases of suspected ADRs reports in order to identify previous published case reports and spontaneous reports about the ADRs reviewed in this paper, and to examine the role of suspected drugs in the pathogenesis of the described adverse reactions. RESULTS: First, we reported a case of tizanidine-induced hemorrhagic cystitis. In the second case report, we presented an episode of asthma exacerbation after taking bimatoprost. Through the review of these two cases, we highlighted some common criticisms of spontaneous reporting systems: under-reporting and false causality attribution. DISCUSSION AND CONCLUSION: Healthcare workers sometimes do not report ADRs because it is challenging to establish with certainty the causal relationship between drug and adverse reaction; however, according to a key principle of pharmacovigilance, it is always better to report even a suspicion to generate an alarm in the interest of protecting public health.

Corticosteroid-related central nervous system side effects.

Corticosteroids have been used since the 50s as anti-inflammatory and immunosuppressive drugs for the treatment of several pathologies such as asthma, allergy, rheumatoid arthritis, and dermatological disorders. Corticosteroids have three principal mechanisms of action: 1) inhibit the synthesis of inflammatory proteins blocking NF-kB, 2) induce the expression of anti-inflammatory proteins by IkB and MAPK phosphatase I, and 3) inhibit 5-lipoxygenase and cyclooxygenase-2. The efficacy of glucocorticoids in alleviating inflammatory disorders results from the pleiotropic effects of the glucocorticoid receptors on multiple signaling pathways. However, they have adverse effects: Growth retardation in children, immunosuppression, hypertension, hyperglycemia, inhibition of wound repair, osteoporosis, metabolic disturbances, glaucoma, and cataracts. Less is known about psychiatric or side effects on central nervous system, as catatonia, decreased concentration, agitation, insomnia, and abnormal behaviors, which are also often underestimated in clinical practice. The aim of this review is to highlight the correlation between the administration of corticosteroids and CNS adverse effects, giving a useful guide for prescribers including a more careful assessment of risk factors and encourage the use of safer doses of this class of drugs.

CB1 agonists, locally applied to the cortico-thalamic circuit of rats with genetic absence epilepsy, reduce epileptic manifestations.

Drugs that modulate the endocannabinoid system and endocannabinoids typically play an anticonvulsant role although some proconvulsant effects have been reported both in humans and animal models. Moreover, no evidence for a role of the cannabinoid system in human absence epilepsy has been found although limited evidence of efficacy in relevant experimental animal models has been documented. This study aims to characterize the role of cannabinoids in specific areas of the cortico-thalamic network involved in oscillations that underlie seizures in a genetic animal model of absence epilepsy, the WAG/Rij rat. We assessed the effects of focal injection of the endogenous cannabinoid, anandamide (AEA), a non-selective CB receptor agonist (WIN55,212) and a selective CB1 receptor antagonist/inverse agonist (SR141716A) into thalamic nuclei and primary somatosensory cortex (S1po) of the cortico-thalamic network. AEA and WIN both reduced absence seizures independently from the brain focal site of infusion while, conversely, rimonabant increased absence seizures but only when focally administered to the ventroposteromedial thalamic nucleus (VPM). These results, together with previous reports, support therapeutic potential for endocannabinoid system modulators in absence epilepsy and highlight that attenuated endocannabinergic function may contribute to the generation and maintenance of seizures. Furthermore, the entire cortico-thalamic network responds to cannabinoid treatment, indicating that in all areas considered, CB receptor activation inhibits the pathological synchronization that subserves absence seizures. In conclusion, our result might be useful for the identification of future drug therapies in absence epilepsy.

Lack of efficacy during the switch from brand to generic allopurinol.

We report for the first time the lack of therapeutic effects after the switch from a brand formulation of allopurinol to a generic one. A 56-year-old man, with a 5 years history of well-treated gout arthropathy with allopurinol (Zyloric(®) 300 mg/die), developed acute gout arthropathy after the switch from the brand formulation of allopurinol to a generic one. Clinical evaluation and laboratory findings confirmed the diagnosis of acute gout arthropathy. Generic formulation of the drug was dismissed and Zyloric(®) was administered with an improvement of both clinical symptoms and laboratory findings. In conclusion, even if generic formulations are considered to have the same effects in comparison to the brand one, more data are necessaries in order to well define their effectiveness and rationale use.

Antiepileptic action of N-palmitoylethanolamine through CB1 and PPAR-α receptor activation in a genetic model of absence epilepsy.

N-palmitoylethanolamine (PEA), an endogenous fatty acid ethanolamide, plays a key role in the regulation of the inflammatory response and pain through, among others, activation of nuclear peroxisome proliferator-activated receptors (PPAR-α). Endogenous cannabinoids play a protective role in several central nervous system (CNS) disorders, particularly those associated with neuronal hyperexcitability. We investigated the effects of PEA and the role of PPAR-α in absence epilepsy using the WAG/Rij rat model. PEA, anandamide (AEA), a PPAR-α antagonist (GW6471) and a synthetic CB1 receptor antagonist/inverse agonist (SR141716) were administered to WAG/Rij rats in order to evaluate the effects on epileptic spike-wave discharges (SWDs) on EEG recordings. We studied also the effects of PEA co-administration with SR141716 and GW6471 and compared these effects with those of AEA to evaluate PEA mechanism of action and focusing on CB1 receptors and PPAR-α. Both PEA and AEA administration significantly decreased SWDs parameters (absence seizures). In contrast, GW6471 was devoid of effects while SR141716 had pro-absence effects. The co-administration of SR141716 with PEA or AEA completely blocked the anti-absence effects of these compounds. GW6471 antagonized PEA's effects whereas it did not modify AEA's effects. Furthermore, we have also measured PEA, AEA and 2-AG (2-arachidonoylglycerol) brain levels identifying significant differences between epileptic and control rats such as decreased PEA levels in both thalamus and cortex that might contribute to absence epilepsy. Our data demonstrate that PEA has anti-absence properties in the WAG/Rij rat model and that such properties depend on PPAR-α and indirect activation of CB1 receptors. This article is part of the Special Issue entitled 'New Targets and Approaches to the Treatment of Epilepsy'.

Does antiepileptogenesis affect sleep in genetic epileptic rats?

Recently it was established that early long lasting treatment with the anti-absence drug ethosuximide (ETX) delays the occurrence of absences and reduces depressive-like symptoms in a genetic model for absence epilepsy, rats of the WAG/Rij strain. Here it is investigated whether anti-epileptogenesis (chronic treatments with ETX for 2 and 4 months) affects REM sleep in this model. Four groups of weaned male WAG/Rij rats were treated with ETX for 4 months, two groups for 2 months (at 2-3 and 4-5 months of age), the fourth group was untreated. Next, the rats were recorded 6 days after the last day of the treatment for 22.5 h. Non-REM sleep and REM sleep parameters and delta power were analyzed in four characteristic and representative hours of the recoding period. Four months treatment with ETX reduced the amount of REM sleep and REM sleep as percentage of total sleep time. Other sleep parameters were not affected by the treatment. Clear differences between the various hours of the light-dark phase in amounts of non-REM and REM sleep and delta power were found, in line with commonly reported circadian sleep patterns. It can be concluded that the reduction of REM sleep is unique for the early and long lasting chronic treatment. The outcomes may explain our earlier finding that a reduction of REM sleep might alleviate depressive like symptoms.

Pharmacovigilance and drug safety 2011 in Calabria (Italy): Adverse events analysis.

BACKGROUND: Pharmacovigilance assesses the safety profile of drugs. Its main aim is the increase of spontaneous reporting of adverse drug reactions (ADRs). The Italian Drug Agency (AIFA; Agenzia Italiana del Farmaco) is financing several projects to the aim of increasing reporting, and in Calabria a Pharmacovigilance Information Centre has been created. MATERIALS AND METHODS: We analyzed the AIFA database relatively to Calabria in the year 2011 and we have analyzed ADRs using descriptive statistics. We have also collected a questionnaire-based interview in order to describe the background knowledge in the field. RESULTS: Regarding the number of AIFA reported ADRs from Calabria, a 38% increase (138 vs. 100) in comparison to 2010 was evidenced. Hospital Doctors represent the main source of signaling (71.7 %). Ketoprofene and the combination amoxicillin/clavulanic acid represent the most frequently reported drugs causing ADRs. Our questionnaires indicated that despite the health professionals have met at least once an ADR only a small percentage of them was reported to the authorities (37%). There is a very good knowledge of the ADR concept and reporting system (90% of interviewed distinguish an ADR and knows how to report it), and there is a strong interest in participating to training courses in the field (95% are interested). CONCLUSIONS: Despite Calabria has had a positive increase in the number of reported ADRs, the total number is very low and the pharmacovigilance culture is far from being achieved in this region.

Vigabatrin has antiepileptogenic and antidepressant effects in an animal model of epilepsy and depression comorbidity.

To evaluate the effects of Vigabatrin (VGB) treatment, on both absence seizure and depressive-like behaviour development in the WAG/Rij rat model of absence seizures. Early long-term treatment with VGB could alter the development of absence pathology, by significantly reducing seizure generation and synchronization in contrast to its pro-absence effects observed after acute or subchronic administration. We have demonstrated the antidepressant effects of a sub-chronic treatment with VGB in both wistar and WAG/Rij rats. In contrast, following an early long-term treatment, VGB antidepressant effects were only observable in WAG/Rij rats. In conclusion, VGB has antiepileptogenic and antidepressant properties in the WAG/Rij rat model despite its pro-absence effects suggesting that epilepsy and depression, in this animal model, are directly related and that seizure development inhibition also reduces the development of depressive behaviour.

Effects of early long-term treatment with antiepileptic drugs on development of seizures and depressive-like behavior in a rat genetic absence epilepsy model.

PURPOSE: Depression is most commonly associated with epilepsy. Recent reports have suggested a putative relationship between seizure development and onset of depressive behavior, whereas others proposed that two clinical entities might represent different neuropathologic aspects of the same neurologic disorder. The WAG/Rij rat absence epilepsy model has also been proposed as a suitable model to test antidepressant drugs. We previously reported on a long-term study of two antiepileptic drugs (AEDs) to assess their protective role in absence epileptogenesis. Here, we examined the effects of long-term treatment with several AEDs on absence seizure development and onset of depressive-like behavior in WAG/Rij rats at different ages, using a forced swimming test (FST). METHODS: Animals were divided into one untreated control group and four test groups, given ethosuximide, levetiracetam, zonisamide, or carbamazepine. Electroencephalography (EEG) readings were recorded at 6.5 months of age. KEY FINDINGS: Ethosuximide-treated animals showed significant reductions in recorded spike-wave discharges (SWDs), and FST immobility time (IT) compared with untreated same age controls. However, zonisamide- and carbamazepine-treated animals had IT values similar to those of controls, but only zonisamide significantly decreased absence seizure development. Carbamazepine increased SWD incidence. Levetiracetam also protected against seizure development, while augmenting IT, suggesting a prodepressive effect. SIGNIFICANCE: Although treatment with ethosuximide, levetiracetam, or zonisamide reduced appearance of SWDs in WAG/Rij rats, this was not generally linked to a reduced onset of depressive characteristics, as assessed by FST. Therefore, expression of depressive-like behavior seems unrelated to seizure control in this model. Some possible alternative explanations for the observed data are discussed.

Preclinical activity profile of α-lactoalbumin, a whey protein rich in tryptophan, in rodent models of seizures and epilepsy.

PURPOSE: To evaluate the potential anticonvulsant activity of α-lactalbumin (ALAC), a whey protein rich in tryptophan (TRP) relative to other large neutral amino acids (LNAAs), in rodent models of seizures and epilepsy. METHODS: The effects of ALAC administered per os were evaluated by standard protocols against audiogenic seizures in Genetic Epilepsy Prone Rats (GEPR-9 rats), maximal electroshock (MES)-induced seizures in rats, pilocarpine-induced seizures in mice, spontaneous chronic seizures in mice exposed to pilocarpine-induced status epilepticus (SE), and absence seizures in WAG/Rij rats. In some models, carbamazepine (CBZ) was included as an active control. Plasma TRP/LNAAs ratios were measured by GC-MS. RESULTS: Single doses of ALAC up to 500 or 6000 mg/kg were devoid of anticonvulsant activity in all models tested. Conversely, 5- and 12-day treatment with ALAC (250-1000 mg/kg/day) in GEPR rats reduced dose-dependently seizure scores and prolonged latency to clonus onset, with full persistence of the effect for up to 12h. ALAC (125-500 mg/kg/day for 15 days) protected against seizures induced by 250 mg/kg pilocarpine, but was less effective against higher pilocarpine doses. Similarly to CBZ, ALAC (125-500 mg/kg/day for 15 days) was also effective against spontaneous seizures in the post-pilocarpine SE model. ALAC (up to 6000 mg/kg/day for 12 days) did not prevent MES-induced seizures, although it reduced the duration of tonic extension at doses between 250 and 1000 mg/kg/day. Absence seizures in WAG/Rij rats were not significantly affected by ALAC. Plasma TRP/LNAAS ratios increased 2- to 3-fold after dosing with ALAC (250 mg/kg/day) for 7 and 14 days, respectively. CONCLUSIONS: ALAC exerts significant protective activity against seizures in animal models, the effect being especially prominent against audiogenic seizures in GEPR-9 rats, seizures induced by low-dose pilocarpine in mice, and spontaneous seizures in mice exposed to pilocarpine-induced SE. This action is likely to be mediated by increased availability of TRP in the brain, with a consequent increase in 5-HT mediated transmission.

Comparison of the antiepileptogenic effects of an early long-term treatment with ethosuximide or levetiracetam in a genetic animal model of absence epilepsy.

PURPOSE: Epilepsy is a heterogeneous syndrome characterized by recurrent, spontaneous seizures; continuous medication is, therefore, necessary, even after the seizures have long been suppressed with antiepileptic drug (AED) treatments. The most disturbing issue is the inability of AEDs to provide a persistent cure, because these compounds generally suppress the occurrence of epileptic seizures without necessarily having antiepileptogenic properties. The aim of our experiments was to determine, in the WAG/Rij model of absence epilepsy, if early long-term treatment with some established antiabsence drugs might prevent the development of seizures, and whether such an effect could be sustained. METHODS: WAG/Rij rats were treated for ∼3.5 months (starting at 1.5 months of age, before seizure onset) with either ethosuximide (ETH; drug of choice for absence epilepsy) or levetiracetam (LEV; a broad-spectrum AED with antiabsence and antiepileptogenic properties). RESULTS: We have demonstrated that both drugs are able to reduce the development of absence seizures, exhibiting antiepileptogenic effects in this specific animal model. DISCUSSION: These findings suggest that absence epilepsy in this strain of rats very likely follows an epileptogenic process during life and that early therapeutic intervention is possible, thereby opening a new area of research for absence epilepsy and AED treatment strategies.

Solution-phase parallel synthesis and evaluation of anticonvulsant activity of N-substituted-3,4-dihydroisoquinoline-2(1H)-carboxamides.

In our previous studies we identified several isoquinoline derivatives displaying potent anticonvulsant effects in different animal models of epilepsy. With the aim to exploit the main structure-activity relationships (SAR) for this class of compounds we planned a solution-phase parallel synthesis (SPPS) of new N-substituted-3,4-dihydroisoquinoline-2(1H)-carboxamides exploring the effect of introduction of different (cyclo)alkyl groups at carboxamide moiety linked to N-2 atom of isoquinoline scaffold. The pharmacological effects were evaluated against audiogenic seizures in DBA/2 mice and, even if some new derivatives were more active than valproate, the designed modifications did not improve the anticonvulsant efficacy with respect to their precursors.

Solution-phase parallel synthesis of novel 1,2,3,4-tetrahydroisoquinoline-1-ones as anticonvulsant agents.

Following our previous studies in the field of anticonvulsant agents, we planned a one-pot solution-phase parallel synthesis (SPPS) of a small library of new 1,2,3,4-tetrahydroisoquinoline derivatives. The activity against audiogenic seizures in DBA/2 mice of the newly synthesized compounds has also been evaluated.